Added Magnesium Ions Research Articles

A bionic composite hydrogel with dual regulatory functions for the osteochondral repair

Due to the avascular nature of cartilage, it is difficult to heal and regenerate spontaneously after injury. At present, tissue engineering has become a promising strategy for repairing damaged cartilage, but the use of seed cells and growth factors is limited. In addition, the importance of mechanical compatibility of scaffold materials is often ignored. In this study, osteochondral scaffold was designed as a bilayer structure with a dense γ-Polyglutamic acid/carboxymethyl chitosan/bacterial cellulose (PGA/CMCS/BC) hydrogel cartilage layer and a porous nano HA-containing PGA/CMCS/BC hydrogel osteogenic layer. In addition, bioactive ions were introduced into the hydrogel scaffold to adjust the mechanical and swelling properties of the material to match the mechanical properties of natural articular cartilage. At the same time, based on the structural characteristics of bone and cartilage, magnesium and copper ions were introduced into the double-layer hydrogel scaffold, respectively, to prepare the cartilage layer and the bone layer, which endowed the material with excellent antibacterial properties and achieved the purpose of the integrated repair of bone and cartilage. The results showed that, after adding magnesium ions, the tensile breaking strength of material was increased from 0.66 MPa to 1.37 MPa，the corresponding compression modulus of the material (strain 0–12%) increased from 0.15 MPa to 0.58 MPa whilst the maximum mass swelling rate decreased from 155% to 75%. The results of in vivo experiments show that the group with bioactive ions had a much better effect on the repair of osteochondral defects, compared with group without bioactive ions, demonstrating such double ion regulation strategy is a very practical strategy for the treatment of osteochondral defects.

Enhanced struvite generation and separation by magnesium anode electrolysis coupled with cathode electrodeposition

Adding magnesium ions (Mg2+) to produce struvite is an important method to recover nitrogen and phosphorus from wastewater. Both the Mg2+ source and subsequent separation of struvite are key factors for the utilization of struvite. In this study, we developed an efficient method to recover nutrient salts from wastewater using sacrificial Mg anodes to generate struvite, with its simultaneous separation through cathode electrodeposition. The anode-released Mg2+ reacted with NH4+-N and PO43−-P in bulk solution to form struvite, which was more intense on the cathode surface due to the relatively higher pH environment from hydrogen evolution, resulting in most of the struvite being deposited on the cathode surface and simultaneously separated out of the bulk solution. Using a cathode with a higher solution-cathode interface area and relatively low current density facilitated struvite deposition. Results showed that under optimal electrolysis condition (5.76 A/m2, pH 8.5, 180 min, and 1.2:1.0 Mg:P), 91% of the undissolved substances as the phosphate precipitation were deposited on the graphite cathode surface, and the proportion of struvite in the deposition reached 41.52%. This study provides a novel electrochemical method for struvite synthesis and separation for the recovery of nitrogen and phosphorus from wastewater.

Ultrahigh permeance of a chemical cross-linked graphene oxide nanofiltration membrane enhanced by cation-π interaction.

Cross-linking with large flexible molecules is a common method to improve the stability and control the interlayer spacing of graphene oxide (GO) membranes, but it still suffers from the limitation of low water flux. Herein, a novel high flux GO membrane was fabricated using a pressure-assisted filtration method, which involved a synergistic chemical cross-linking of divalent magnesium ions and 1,6-hexanediamine (HDA) on a polyethersulfone (PES) support. The membrane cross-linked with magnesium ions and HDA (GOHDA–Mg2+) exhibited a high water flux up to 144 L m−2 h−1 bar−1, about 7 times more than that of cross-linked GO membranes without adding magnesium ions (GOHDA), while keeping excellent rejection performance. The GOHDA–Mg2+ membrane also showed an outstanding stability in water for a long time. The effects of magnesium ions on the GOHDA–Mg2+ membrane were analyzed using several characterization methods, including Fourier transform infrared spectroscopy (FT-IR), scanning electron microscopy (SEM), X-ray photoelectron spectroscopy (XPS) and X-ray diffraction (XRD). The results indicated that magnesium ions not only promoted reasonable cross-linking, but also improved the stacking of GO sheets to give lower mass transfer resistance channels for water transport in the membranes, resulting in the ultrahigh permeance of the GO membranes.

Magnesium ions increase the activity of Bacillus deramificans pullulanase expressed by Brevibacillus choshinensis.

Addition of MgCl2 to the culture medium has been found to dramatically increase the activity of Bacillus deramificans pullulanase expressed by Brevibacillus choshinensis. The specific activity of the pullulanase obtained from medium supplemented with MgCl2 was also higher than that obtained in culture medium without added magnesium ions. In this work, the mechanism of this increase was studied. When cultured in medium without added magnesium ions, B. choshinensis mainly produced a thermolabile, inactive form of pullulanase. The addition of magnesium ions led to the production of a thermostable, active form of pullulanase. Circular dichroism assays revealed considerable differences in secondary structure between the active and inactive pullulanase forms. Transmission electron microscopy suggested that magnesium ion addition inhibits the shedding of cell wall protein (HWP) layers from the cell surface. Quantitative real-time PCR showed that magnesium ion addition represses transcription of HWP. Because the pullulanase gene and HWP have identical promoters, pullulanase gene transcription was also inhibited. These results suggest that when pullulanase is expressed slowly, it tends to fold into an active form.

Design model for magnesium ions re-mineralization of desalinated water by dissolution of magnesia pellets

Re-mineralization of desalinated water with magnesium ions is currently under consideration. A simple easily controlled technique for adding magnesium ions to desalinated water consists of dissolution of magnesia pellets in a packed bed by feed water slightly acidified with either carbon dioxide or sulfuric acid. A design model is presented describing magnesia pellets dissolution in a fixed bed as a mass transfer controlled process. The model is confirmed by experimental data covering a range of acid concentrations and contact times. A striking difference was observed between dissolution with the weak carbonic acid and with the strong sulfuric acid. Dissolution of magnesia with dilute sulfuric acid of 1 to 5mM at very short contact times around 2min provided virtual equilibrium concentration of magnesium ions whereas the magnesium ion concentrations obtained with CO2 at the same acid concentrations and contact times were far from equilibrium. Mass transfer coefficients extracted from the dissolution data of the weak carbonic acid were in substantial agreement with literature data whereas coefficients derived from the strong sulfuric acid dissolution data were about 5–6 times higher, probably due to an increased pellet surface generated by a strong acid attack.

Facile synthesis of calcium carbonate with an absolutely pure crystal form using 1-butyl-3-methylimidazolium dodecyl sulfate as the modifier

Calcium carbonate (CaCO3) nanocrystals with controllable polymorph and morphology have been successfully synthesized with the aid of an effective control agent, a halogen-free, low-cost ionic liquid surfactant, 1-butyl-3-methylimidazolium dodecylsulfate ([C4mim][C12SO4]) in a supersaturated aqueous solution. For the first time, facile preparation of pure lens-like vaterite, sheet-like calcite, and peanut-like aragonite was all achieved in the [C4mim][C12SO4] aqueous solution through changing the concentration, temperature, and initial pH value and adding magnesium ions. Washed by water and ethanol, all the aggregates were free of [C4mim][C12SO4] and can be stable at least 1 month in air. The crystal form of the aggregates changed from pure calcite to pure vaterite at room temperature only through increasing [C4mim][C12SO4] concentration. Formation of the ordered CaCO3 structures is mainly ascribed to the aggregation of the primary nanoparticles whose formation mechanism is related to the change of supersaturation. This study can provide a facile and environment-friendly method to fabricate CaCO3 crystal aggregates with various morphologies and polymorphs and can be used for large-scale industrial production and biomimetic synthesis.

Potential applications of quarry dolomite for post treatment of desalinated water

Recent WHO publications recommend a minimum concentration of 10 mg Mg/L in desalinated water. Dolomite dissolution was investigated as means of adding magnesium ions to desalinated water at the post treatment stage, in addition to Ca2+ ions and carbonate alkalinity. The results show that dolomite dissolution per se is not feasible for post treatment purposes, because dolomite stops dissolving at a relatively low pH, rendering the alkalinity and Calcium Carbonate Precipitation Potential values very low and negative, respectively. To overcome this problem three combined dolomite-calcite dissolution alternatives were investigated. The results show that the most promising method is to dissolve dolomite and then use the CO2(aq)-rich effluent to further dissolve calcite. Applying such in-series dissolution, it is possible to produce water with the following quality criteria: alkalinity = 75 mg/L as CaCO3, [Mg2+] = 12.4 mg/L, [Ca2+] = 120 mg/L as CaCO3, pH 8.17. However, the resultant total hardness value is high (170 mg/L as CaCO3). The operational costs of this alternative were approximated at 0.042 $ m-3 product water.

Protein Phosphatase 2B of Saccharomyces Cerevisiae is Required for Tolerance to Manganese, in Blocking the Entry of ions into the Cells

The role of protein phosphatase 2B (PP2B/calcineurin) of Saccharomyces cerevisiae in the tolerance to divalent cations was investigated. PP2B-deficient mutants were found to be sensitive to MnCl2, but not to ZnCl2, CuCl2, NiCl2 and CoCl2. By measuring both manganese uptake and its efflux, it was found that the sensitivity of the mutant cells was due to an increase in manganese uptake and that the wild-type cells were able to prevent manganese entry into the cells, rather than export it in a more efficient manner. In the presence of the immunosuppressant FK506, the behavior of wild-type cells became similar to that of PP2B mutants. Out of various divalent cations tested, externally added magnesium ions were able to block manganese uptake in both wild-type and PP2B mutant strains.

Entrapment in E. coli improves the operational stability of recombinant β-glycosidase CelB from Pyrococcus furiosus and facilitates biocatalyst recovery

beta-Glycosidase CelB from the hyperthermophilic archaeon Pyrococcus furiosus is a versatile biocatalyst that has been used for the hydrolysis and synthesis of beta-d-glycosidic compounds at high temperatures and in non-conventional solvents. In spite of its outstanding thermal stability, CelB is prone to inactivation in the presence of reducing sugars and through recirculation in loop enzyme reactors. Entrapment into E. coli cells was used here to improve the stability of recombinant CelB under conditions promoting strong inactivation. Glutardialdehyde-mediated protein cross-linking or rigidification of the cell membrane by adding magnesium ions was required to prevent release of CelB from within the cell into the bulk solution. In the presence of 1M glucose or when applying recirculation rates of 2.6 min(-1), the entrapped enzyme was around two-fold more stable at 80 degrees C than free CelB. The significance of the stabilisation was attenuated by the decrease in CelB initial activity which was due to cross-linking and glutardialdehyde concentration-dependent. Entrapment facilitated downstream processing of CelB and biocatalyst recovery in repeated batchwise conversions of lactose at elevated temperatures.

Molecular Level Probing of Preferential Hydration and Its Modulation by Osmolytes through the Use of Pyranine Complexed to Hemoglobin

Two spectroscopic probes are used to expose molecular level changes in hydration shell water interactions that directly relate to such issues as preferential hydration and protein stability. The major focus of the present study is on the use of pyranine (HPT) fluorescence to probe as a function of added osmolytes (PEG, urea, trehalose, and magnesium), the extent to which glycerol is preferentially excluded from the hydration shell of free HPT and HPT localized in the diphosphoglycerate (DPG) binding site of hemoglobin in both solution and in Sol-Gel matrices. The pyranine study is complemented by the use of vibronic side band luminescence from the gadolinium cation that directly exposes the changes in hydrogen bonding between first and second shell waters as a function of added osmolytes. Together the results form the basis for a water partitioning model that can account for both preferential hydration and water/osmolyte-mediated conformational changes in protein structure.

Electron Transfer to Photosystem 1 from Spinach Plastocyanin Mutated in the Small Acidic Patch: Ionic Strength Dependence of Kinetics and Comparison of Mechanistic Models

A set of plastocyanin (Pc) mutants, probing the small acidic patch (Glu59, Glu60, and Asp61) and a nearby residue, Gln88, has been constructed to provide further insight into the electron transfer process between Pc and photosystem 1. The negatively charged residues were changed into their neutral counterparts or to a positive lysine. All mutant proteins exhibited electron transfer kinetics qualitatively similar to those of the wild type protein over a wide range of Pc concentrations. The kinetics were slightly faster for the Gln88Lys mutant, while they were significantly slower for the Glu59Lys mutant. The data were analyzed with two different models: one involving a conformational change of the Pc-photosystem 1 complex that precedes the electron transfer step (assumed to be irreversible) [Bottin, H., and Mathis, P. (1985) Biochemistry 24, 6453-6460] and another where no conformational change occurs, the electron transfer step is reversible, and dissociation of products is explicitly taken into account [Drepper, F., Hippler, M., Nitschke, W., and Haehnel, W. (1996) Biochemistry 35, 1282-1295]. Both models can account for the observed kinetics in the limits of low and high Pc concentrations. To discriminate between the models, the effects of added magnesium ions on the kinetics were investigated. At a high Pc concentration (0.7 mM), the ionic strength dependence was found to be consistent with the model involving a conformational change but not with the model where the electron transfer is reversible. One residue in the small acidic patch, Glu60, seems to be responsible for the major part of the ionic strength dependence of the kinetics.

Operator interactions by the Bacillus subtilis arginine repressor/activator, AhrC: novel positioning and DNA-mediated assembly of a transcriptional activator at catabolic sites.

We have previously reported the initial characterization of a catabolic operator site (O[rocA]) for the Bacillus subtilis arginine repressor/activator protein AhrC. Here, we present the characterization by gel retardation and DNase I footprinting of both O(rocA) and a second catabolic operator site, O(rocD). Both operator sites encompass a single recognition site, an ARG box, located immediately upstream of the transcriptional start points, a unique positioning for a transcriptional activator protein. Although there is considerable sequence homology between the two catabolic operator sites, they vary significantly, around twofold, in their apparent affinities for the protein (K'd approximately 90 nM for O[rocA] and approximtaely 190nM for O[rocD]). This difference may result from the lower match to the ARG box consensus of the O(rocD) site. Both catabolic operators show evidence for co-operative binding with respect to protein concentration. Determination of the sequences of two AhrC catabolic operator sites, in combination with the three such biosynthetic sites, has allowed the derivation of an improved B. subtilis ARG box consensus sequence, CATGAATAAAAATg/tCAAg/t. This is not identical to the Escherichia coli consensus operator for the AhrC homologue, ArgR, which may explain the only partial cross-functioning of these proteins in vivo. The O(rocA) site is adjacent to a sharp, stable bend located 5' to the catabolic operator. Circular permutation analysis has been used to determine the relative angle of bend (approximately 50 degrees), its location and the effect of adding magnesium ions and/or AhrC protein. Protein binding increases the relative bend angle to approximately 85 degrees. Bending is shown to be associated with a number of A-tracts in the upstream sequence. However, altering the phasing of the A-tracts has little effect on the affinity for AhrC. Truncation and competition experiments have been used to investigate the possible role of sequences flanking the operator on affinity. Very surprisingly, the affinity of the O(rocA) site appears to increase in the presence of excess, specific competitor fragment, i.e. the system shows anti-competitive effects. Competition is restored at high molar excesses of specific fragment over the protein. We propose a novel model for the assembly of a higher affinity form of AhrC at operator sites that is consistent with both the apparent co-operativity of binding and the anti-competitive effects. These data suggest that the molecular interactions occurring between the prokaryotic arginine-regulatory proteins and their operators may be more complex than is generally appreciated.

Rapid-reaction analysis of plasmid DNA cleavage by the EcoRV restriction endonuclease.

Rapid-reaction methods have been used previously to identify intermediates in the reaction of the EcoRV restriction endonuclease on oligonucleotide substrates. In this study, the pathway on macromolecular DNA was elucidated by using the quench-flow method to analyze EcoRV reactions on a plasmid with one recognition site. Some reactions were carried out by first allowing the EcoRV enzyme to bind nonspecifically to the DNA and then initiating DNA cleavage by adding magnesium ions. The subsequent transfer of the enzyme from nonspecific to specific sites was extremely rapid, at a random walk rate of at least 5 x 10(5) base pairs per second. The two strands of the DNA at the EcoRV recognition site were then cleaved sequentially, at rates that were faster than the turnover number of the enzyme. The rates recorded for the cleavage steps were direct measurements of phosphodiester hydrolysis, while the turnover is limited by the dissociation of the product cleaved in both strands. Other reactions were initiated by adding EcoRV and MgCl2 to the DNA: these revealed not only the processes observed in reactions starting from DNA-bound enzyme but also the bimolecular association of the protein with the plasmid. The association rate was limited by diffusion but its rate constant, 1.2 x 10(8) M(-1) s(-1), was unusually small for the binding of a protein to DNA. The slowness of this diffusion-controlled process may be due to a rapid oscillation of the protein between closed and open conformations, with only the open form capable of binding DNA.

Yeast Mitochondrial F1-ATPase-Effects of Metal-Ions

The effects of a series of metal ions (M) on the velocity (V) of the reaction catalyzed by the yeast mitochondrial F1-ATPase were investigated in N-tris(hydroxymethyl)methyl-2-aminoethanesulfonic acid and Imidazole buffers (pH 7.45), with total concentrations of ATP ranging from 2 to 8 mM. The biphasic metal ion activation/inhibition effects, observed with a constant total amount of ATP, were found to be consistent with a simple empirical kinetic equation, V1/V = 1 + K1/[M] + [M]/K2, where V1, K1, and K2 are empirical parameters and [M] the concentrations of metal ions not complexed with the ATP. Three alternative kinetic equations, involving the concentrations of either ATP or its metal chelate, consistent with this empirical equation, were shown to be invalid since changing the total amount of ATP did not affect the empirical reaction parameters V1, K1, and K2. The fact that changing the total ATP concentration failed to change the empirical reaction parameters suggests that the inhibition by an excess of ATP, which is observed with a constant total metal ion concentration, is due to chelation of free metal ions that are required for the ATP hydrolysis. When the total amount of added ATP was in excess of that of added magnesium ions it was found that calcium, strontium, and aluminum ions, which liberate free magnesium from the Mg-ATP chelate by complexing the ATP, all activated. These activations apparently confirm the fact that the ATPase requires a free magnesium ion as a cofactor for the hydrolysis of ATP.

NADH Oxidase Activity of Plasma Membranes of Soybean Hypocotyls Is Activated by Guanine Nucleotides.

The activity of an auxin-stimulated NADH oxidase of the plasma membrane of hypocotyls of etiolated soybean (Glycine max Merr.) seedlings responded to guanine and other nucleotides, but in a manner that differed from that of enzymes coupled to the classic trimeric and low molecular weight monomeric guanine nucleotide-binding proteins (G proteins). In the presence and absence of either auxin or divalent ions, both GTP and GDP as well as guanosine-5[prime]-O-(3-thiotriphosphate) (GTP-[gamma]-S) and other nucleoside di- and triphosphates stimulated the oxidase activity over the range 10 [mu]M to 1 mM. GTP and GTP-[gamma]-S stimulated the activity at 10 nM in the absence of added magnesium and at 1 nM in the presence of added magnesium ions. Other nucleotides stimulated at 100 nM and above. The NADH oxidase was stimulated by 10 [mu]M mastoparan and by 40 [mu]M aluminum fluoride. Neither cholera nor pertussis toxins, tested at a concentration sufficient to block mammalian G protein function, inhibited the activity. Guanosine 5[prime]-O-(2-thiodi-phosphate) (GDP-[beta]-S) did not stimulate activity, suggesting that the stimulation in response to GDP may be mediated by a plasma membrane nucleoside diphosphate kinase through conversion of GDP to GTP. Auxin stimulation of the NADH oxidase was unaffected by nucleotides at either high or low nucleotide concentrations in the absence of added divalent ions. However, pretreatment of plasma membranes with auxin increased the apparent affinity for nucleotide binding. This increased affinity, however, appeared not to be the mechanism of auxin stimulation of the oxidase, since auxin stimulation was similar with or without low concentrations of guanine nucleotides. The stimulation by nucleotides was observed after incubating the membranes with 0.1% Triton X-100 prior to assay. The results suggest a role of guanine (and other) nucleotides in the regulation of plasma membrane NADH oxidase that differs from the interactions with G proteins commonly described for animal models.

Electrooptical studies on proton-binding and -release of bacteriorhodopsin

Electric field induced pH changes of purple membrane suspensions were investigated in the pH range from 4.1 to 7.6 by measuring the absorbance change of pH indicators. In connection with the photocycle and proton pump ability, three different states of bacteriorhodopsin were used: (1) the native purple bacteriorhodopsin (magnesium and calcium ions are bound, the M intermediate exists in the photocycle and protons are pumped), (2) the cation-depleted blue bacteriorhodopsin (no M intermediate), and (3) the regenerated purple bacteriorhodopsin which is produced either by raising the pH or by adding magnesium ions (the M intermediate exists). In the native purple bacteriorhodopsin there are, at least, two types of proton binding sites: one releases protons and the other takes up protons in the presence of the electric field. On the other hand, blue bacteriorhodopsin and the regenerated purple bacteriorhodopsin (pH increase) show neither proton release nor proton uptake. When magnesium ions are added to the suspensions, the field-induced pH change is observed again. Thus, the stability of proton binding depends strongly on the state of bacteriorhodopsin and differences in proton binding are likely to be related to differences in proton pump activity. Furthermore, it is suggested that the appearance of the M intermediate and proton pumping are not necessarily related.

Effects of phencyclidine, SKF 10,047 and related psychotomimetic agents on N-methyl-D-aspartate receptor mediated synaptic responses in rat hippocampal slices.

The effects of representative drugs from three classes of psychotomimetic compounds (arylcyclohexylamines, benzomorphan opioids and dioxolanes) have been examined on synaptic transmission at an identified monosynaptic pathway in rat hippocampal slices. The compounds tested were phencyclidine (PCP) and ketamine, the racemate and isomers of SKF 10,047 (N-allylnormetazocine), and the isomers of dioxadrol (dexoxadrol and levoxadrol). In the absence of added magnesium ions (Mg) in the perfusion medium low frequency stimulation of the Schaffer collateral-commissural pathway evoked a burst of population spikes in the CA1 cell body region. The secondary components of this response could be abolished by the selective N-methyl-D-aspartate (NMDA) antagonist D-2-amino-5-phosphonovalerate (APV). PCP (1 microM) or ketamine (10 microM) selectively blocked the secondary components of the synaptic response. The effect of PCP was neither mimicked nor prevented by hexamethonium and atropine, phentolamine and propranolol, or clonidine and was therefore unlikely to involve cholinergic or adrenergic neurotransmitter systems. The sigma opiate, (+/-)-SKF 10,047 (10 microM) also abolished selectively the secondary components of the synaptic response. There was no apparent difference between the potency of the stereoisomers of this compound. The action of (+/-)-SKF 10,047 was not affected by either naloxone or haloperidol, indicating that this effect did not involve opioid receptors or the haloperidol-sensitive sigma site. Dexoxadrol (10 microM), but not levoxadrol (10 microM), also selectively blocked the secondary components of the synaptic response. It is concluded that these psychotomimetic agents can block an NMDA receptor-mediated component of synaptic transmission in the hippocampus and that this effect is mediated by a specific PCP/sigma site.

Interactions Between Rolling Oil Emulsions and Aluminum Alloy Surfaces

In oil/water/solid contact angle experiments, aluminum alloy surfaces with a high percentage of MgO in the oxide layer were wetted by naphthenic oil and naphthenic oil containing 1 percent oleic acid. The opposite behavior, poor wetting, was observed on alloy surfaces of almost pure aluminum oxide. On the latter surfaces, however, oil wetting was increased by decreasing the pH and by adding magnesium ions to the water being displaced. The same trends in wetting were observed in interactions between pure oxide powders and a commercial rolling emulsion containing naphthenic oil and oleic acid. The implications of this behavior to process lubrication and surface quality in aluminum rolling are discussed. Presented at the 38th Annual Meeting in Houston, Texas, April 24–28, 1983

Cellular levels of the prophage λ and 434 repressors

As a prerequisite to a quantitative study of the inactivation of phage repressors in vivo (Bailone et al., 1979), the cellular concentrations of the bacteriophage λ and 434 repressors have been measured in bacteria with varying repressor levels. Using the DNA-binding assay we have determined the conditions for optimal repressor titration. The sensitivity of the λ repressor assay was increased by adding magnesium ions to the binding mixture; this procedure was without effect on the titration of the 434 repressor. The measures of the cellular repressor concentrations varied with the method of cell disruption. The cellular concentration of λ repressor, about 140 active repressor molecules per monolysogen, was relatively constant under specific cultural conditions. The repressor concentration increased with the number of cI gene copies but not in direct proportion. The 434 repressor concentration, hardly detectable in extracts of lysogens carrying an imm434 prophage, was greatly enhanced in bacteria carrying the newly constructed plasmid pGY101, that encodes the 434 cI gene. The cellular repressor level produced by 434 is lower than that produced by λ: this indicates that the maintenance of the prophage state is ensured by a relatively small number of repressor molecules binding tightly to the operator sites.

Activation of the alternative pathway of complement by antiserum to factor B.

Monospecific rabbit and goat antisera to human complement proteins and human immunoglobulins were tested for their ability to activate the alternative complement pathway. This activation was detected by two methods where classical pathway activation was blocked with EGTA and alternative pathway activation was promoted with added magnesium ions. These two methods consisted of lysis of GSHE and conversion of factor B into split products. C1q-depleted serum was used in a third assay system. Only antiserum to human factor B was able to activate the alternative pathway in the various systems used. None of the other anticomplement sera showed such activity. When antiserum to factor B was fractionated by ammonium sulfate and column chromatography, activation of the alternative pathway was found in the IgG fraction, and this activity was completely removed by absorption with purified factor B but not with other purified complement components.